MOLECULAR REGULATION OF IMMUNOGLOBULIN GENE EXPRESSION

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The experiments proposed here concern the relationship between gene organization and gene expression as it affects the immunoglobulin heavy chain (Igh) gene family, and neighboring genes and gene families on mouse chromosome 12. To construct detailed linkage and physical maps of the chromosome, chromosome-specific DNA fragments isolated from an interspecies somatic cell hybrid will be used to define restriction fragment length polymorphisms and to analyze hybrids carrying subchromosomal fragments. Chromosome "hopping" procedures will be used to characterize the region between the proto-oncogene c-Fos and Igh. To identify transcriptionally active loci in the chromosome, difference hybridization techniques will be used to isolate chromosome-specific, tisssue-specific cDNAs. The expression of these during the development of the immune system will be analyzed by in situ hybridization. To determine the structural basis of the extensive DNA-level polymorphism found in previous experiments, allelic forms of these loci will be cloned and studied by restriction mapping, heteroduplex analysis, and, where necessary, sequence analysis, in order to determine whether particular classes of events (transpositions, non-homologous recombinations) are responsible for the observed diversity. Beyond the insight that these experiments should provide into the genetic organization of mouse chromosome 12 and functional consequences of that organization, the work should help to define strategies more generally for the "mid-level" analysis of mammalian gene organization.